LEIGH E. CHADWICK 51 



the likelihood that unidentified inhibitory components of the crude prepa- 

 rations obscure the true temperature relationships of the enzyme-substrate 

 reaction, there remains the chance that the heats and entropies of activa- 

 tion of the several steps represented by ki , k2 and k^ of reaction 1 might 

 be so balanced as to produce a direct proportionality l)et\veen hydrolysis 

 rate and temperature over a 50° range, although no fully comparable ex- 

 am]ile seems to have been reported. Both possibilities are of course almost 

 wholly speculative in the present state of our knowledge of the system; and 

 experimentally only an attempt to establish or eliminate a role of inhibitory 

 iminirities would seem to be feasible within the limitations of current tech- 

 niques. 



At the same time, one may wonder whether the temperature response of 

 fly head ChE and other preparations with similar properties might not be 

 explained more simply by supposing that the rate limiting process in not 

 one that involves the thermodynamic aspects of the enzyme-substrate re- 

 action, but rather some other type of energy barrier whose height is in- 

 versely proportional to temperature. Such a suggestion is by no means new. 

 Galehr and Plattner (17) concluded, on the basis of their measurements of 

 Qio values of only 1.3 for human blood ChE, that a physical process was 

 concerned ; and they proposed that diffusion of ACh to active sites on the 

 enzyme was the limiting factor. More than the low temperature coefficient, 

 which they stressed, the direct ratio between activity and temperature 

 favors the notion that the rate might be regulated by diffusion. However, 

 attempts to test this hypothesis by altering the viscosity of the medium 

 were made by Kodera (28) , with inconclusive results. We too have reported 

 that, although the rate of hydrolysis is slow^ed in viscous solutions, it is not 

 directly proportional to viscosity in tests made in solutions of sucrose or 

 glycerol ( 11 ) ; but of course w^e do not know what other complications the 

 addition of these substances in high concentrations may have introduced. 

 Still, acceptable evidence that diffusion is the limiting factor has not 

 been provided; and one must remember too that Davies' (15) data with 

 partly purified preparations (fig. 2) and possibly those of Ormerod (34) 

 with benzoylcholine derivatives fit an exponential formulation, even though 

 most other observations do not. 



Thus it seems that no present hypothesis accounts satisfactorily for all 

 the observations, although various explanations are conceivable wuthin the 

 framework of existing theory. Only further comparative work will resolve 

 the question. The problem is both basically important and common to all 

 the ChE's, from whatever source, so that a solution of the dilemma with 

 any one of these enzymes would shed light on the properties of all of them, 

 and perhaps upon unrelated systems as well. 



